I think you are seeing the absorbance due to oxidized DTT---I can't
remember the wavelength maximum for that offhand, but it definitely
absorbs strongly in the far UV (the reduced form has far less
absorbance).
John Philo
Alliance Protein Laboratories
"B.H.Kang" wrote:
>> I prepared the protein with histidine tag in bacterial cell and purified
> with Ni resin.
> For the quantitation of my protein, I check the UV absorption.
> Surprisingly, the maximun absorbance was at 260 nm instead of 280 nm
> and the absorbance was slightly increased comparing the real concentration
> of the protein.
> Is this usual case?
> I think that my protein has no significant nucleic acid contamination.
> And the buffer contains 1 mM DTT,
> The sample buffer was used for baseline of UV absorbance.
>> I didn't know the reason...
>> Thanks and happy new year...!